The aim of the proposed research is to study the nature, distribution, and dynamics of cell surface glycoproteins. First, the functionally and topographically distinct domains of the rat liver parenchymal cell surface will be isolated and purified. A new approach will be used utilizing ligands (antibody or asialo-glycoprotein) which bind specifically to membrane proteins present in only one domain of the cell. The enzymes and glycoproteins of the three plasma membrane fractions will be analysed and compared to establish whether there is extensive or minimal overlap in the molecular composition of the regions. Proteins will be characterized as to number, size, net charge, and carbohydrate content by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectric focusing and lectin labeling in gels, both alone and in combinations. Finally, a limited number of glycoproteins from each domain will be selected (based on abundance, unambiguous assignment as an integral membrane component and presence in intracellular compartments) and kinetic experiments will be conducted in order to trace the intracellular biogenetic, recycling, and degradative pathways of the cell surface glycoproteins. Different labeling protocols (i.e., metabolic labels (amino acids, monosaccharides), exogenous reagents (125I-lactoperoxidase) will be used in isolated, perfused rat liver, followed by subcellular fractionation at various times and analysis of the rates at which compartments (and selected components of them) are labeled. Then the distribution and dynamics of cell surface glycoproteins will be studied in isolated hepatocytes where cell polarity has been perturbed. The liver will be enzymatically dissociated and morphologic techniques (immunofluorescence, immuno-electron microscopy) used to assess the surface redistribution of selected components in single cells. The dynamic properties (bio-genesis, recycling, degradation) of these components in isolated hepatocytes versus intact liver (polarized cells) will be studied using a kinetic approach and subcellular fractionation. Finally, hepatocytes will be cultured under a variety of conditions in order to study the internal and external influences on the re-establishment of cell polarity.